Constitutive stimulation of vascular smooth muscle cells by Angiotensin II derived from an adenovirus encoding a fulin‐cleavable fusion membrane protein

Abstract

Angiotensin II (AngII) plays critical roles in cardiovascular pathophysiology. Acute stimulation of cultured cells by synthetic AngII has been utilized to elucidate the signal transduction mechanisms of the peptide. However, such treatment may not mimic disease conditions where the circulating and/or tissue renin‐angiotensin systems are chronically upregulated to produce AngII consistently. To fill the gap between acute stimulation of cell cultures and chronic stimulation in vivo, and to obtain clinically relevant signaling information from cell cultures, we have made an adenovirus vector encoding a fulin‐cleavable AngII fusion protein. Acute stimulation of vascular smooth muscle cells (VSMCs) with synthetic AngII (100 nM) in cultured medium showed the peptide had a half life of less than 1 hour. In contrast infection of VSMCs with the AngII fusion virus showed a time and dose dependent production of AngII (peaking at 300 pM at 250 moi) in the medium as early as 2 days and up to 7 days post‐infection. Consistent with the peptides function in VSMCs, the AngII fusion virus increased protein content and cell volume, but not cell number and stimulated ERK1/2 activation and EGR‐1 induction. In conclusion, application of the AngII adenovirus to cultured cells will be useful to elucidate molecular and signaling mechanisms of cardiovascular diseases associated with enhanced AngII production.